Alpha-amylase

Abstract

This invention relates to an alpha-amylase, a process for its preparation and the use of the amylase. The invention relates to a newly identified polynucleotide sequence from Alicyclobacillus pohliae comprising a gene that encodes the alpha-amylase enzyme. The invention features the full-length coding sequence of the gene as well as the amino acid sequence of the full-length functional protein of the gene. The invention also relates to methods of using these proteins in industrial processes, for example in food industry, such as the baking industry. Also included in the invention are cells transformed with a polynucleotide according to the invention suitable for producing these proteins and cells.

The invention claimed is: 
     
       1. An isolated polynucleotide comprising:
 (a) SEQ ID NO:1 or SEQ ID NO:3; or 
 (b) nucleotides 100 to 2157 of SEQ ID NO:1; or 
 (c) nucleotides 100 to 2157 of SEQ ID NO:3; or 
 (d) a polynucleotide sequence encoding a polypeptide comprising SEQ ID NO:2; or 
 (e) a polynucleotide sequence encoding a polypeptide comprising amino acids 34 to 719 of SEQ ID NO:2. 
 
     
     
       2. The isolated polynucleotide according to  claim 1 , wherein the polynucleotide is produced by the method comprising:
 (a) obtaining the genome of  Alicyclobacillus pohliae  NCIMB14276; 
 (b) performing sequence analysis on said genome, thereby identifying said polynucleotide in said genome; and 
 (c) isolating said polynucleotide. 
 
     
     
       3. A vector comprising the isolated polynucleotide according to  claim 1 . 
     
     
       4. The vector according to  claim 3  which is an expression vector, wherein the polynucleotide sequence is operably linked with at least one regulatory sequence allowing for expression of the polynucleotide sequence in a suitable host cell. 
     
     
       5. A method for manufacturing the polynucleotide according to  claim 1  comprising culturing an isolated host cell transformed with said polynucleotide and isolating said polynucleotide from said host cell. 
     
     
       6. An isolated polypeptide comprising:
 (a) amino acids 34 to 719 of SEQ ID NO:2; 
 (b) an amino acid sequence encoded by a polynucleotide comprising nucleotides 100 to 2157 of SEQ ID NO:1; or 
 (c) an amino acid sequence encoded by a polynucleotide comprising nucleotides 100 to 2157 of SEQ ID NO:3. 
 
     
     
       7. A method of manufacturing the polypeptide according to  claim 6  comprising cultivating an isolated host cell comprising a vector, the vector comprising a polynucleotide comprising:
 (a) SEQ ID NO:1 or SEQ ID NO:3; or 
 (b) nucleotides 100 to 2157 of SEQ ID NO:1; or 
 (c) nucleotides 100 to 2157 of SEQ ID NO:3; or 
 (d) a polynucleotide sequence encoding a polypeptide comprising SEQ ID NO:2; or 
 (e) a polynucleotide sequence encoding a polypeptide comprising amino acids 34 to 719 of SEQ ID NO:2, 
 under conditions which allow for expression of the vector and, optionally, recovering the encoded polypeptide from the cell or culture medium. 
 
     
     
       8. A method of manufacturing the polypeptide according to  claim 6  comprising cultivating an isolated host cell comprising a polynucleotide, said polynucleotide comprising:
 (a) SEQ ID NO:1 or SEQ ID NO:3; or 
 (b) nucleotides 100 to 2157 of SEQ ID NO:1; or 
 (c) nucleotides 100 to 2157 of SEQ ID NO:3; or 
 (d) a polynucleotide sequence encoding a polypeptide comprising SEQ ID NO:2; or 
 (e) a polynucleotide sequence encoding a polypeptide comprising amino acids 34 to 719 of SEQ ID NO:2, 
 under conditions which allow for expression of the polynucleotide and, optionally, recovering the encoded polypeptide from the cell or culture medium. 
 
     
     
       9. An enzyme composition comprising the polypeptide according to  claim 6  and one or more components selected from the group consisting of: milk powder, gluten, granulated fat, an additional enzyme, an amino acid, a salt, an oxidant, a reducing agent, an emulsifier, a gum, a flavour, an acid, a starch, a modified starch, gluten, a humectant, and a preservative. 
     
     
       10. An enzyme composition according to  claim 9 , wherein the additional enzyme is a lipolytic enzyme. 
     
     
       11. An enzyme composition according to  claim 10 , wherein the lipolytic enzyme is a galactolipase. 
     
     
       12. An enzyme composition according to  claim 10 , wherein the lipolytic enzyme is a phospholipase. 
     
     
       13. An enzyme composition according to  claim 10 , wherein the lipolytic enzyme is an enzyme having both phospholipase and galactolipase activity. 
     
     
       14. A method to prepare a dough comprising combining the polypeptide according to  claim 6  and at least one component selected from the group consisting of: flour, egg, water, salt, sugar, a flavour, a fat, baker's yeast, a chemical leavening system, milk, an oxidant, a reducing agent, an emulsifier, a gum, an acid, starch, modified starch, gluten, an humectant, and a preservative. 
     
     
       15. A method to prepare a dough comprising combining the enzyme composition according to  claim 9  and at least one component selected from the group consisting of:
 flour, egg, water, salt, sugar, a flavour, a fat, baker's yeast, a chemical leavening system, milk, an oxidant, a reducing agent, an emulsifier, a gum, an acid, starch, modified starch, gluten, an humectant, and a preservative. 
 
     
     
       16. A dough comprising the polypeptide according to  claim 6 . 
     
     
       17. A method to prepare a baked product comprising baking a dough comprising the polypeptide according to  claim 6 . 
     
     
       18. A method to prepare a baked product comprising baking a dough comprising the composition according to  claim 9 . 
     
     
       19. A method according to  claim 17 , wherein the baked product is bread or cake. 
     
     
       20. A method according to  claim 18 , wherein the baked product is bread or cake. 
     
     
       21. The isolated polypeptide of  claim 6 , wherein the polypeptide is produced by the method comprising:
 (a) obtaining the genome of  Alicyclobacillus pohliae  NCIMB14276; 
 (b) performing sequence analysis on said genome, thereby identifying in said genome a polynucleotide encoding said polypeptide; 
 (c) isolating said polynucleotide; 
 (d) expressing said polypeptide from said polynucleotide; and 
 (e) isolating said polypeptide.